CN113234657A - 一种无血清细胞培养基的应用 - Google Patents

一种无血清细胞培养基的应用 Download PDF

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CN113234657A
CN113234657A CN202110507226.0A CN202110507226A CN113234657A CN 113234657 A CN113234657 A CN 113234657A CN 202110507226 A CN202110507226 A CN 202110507226A CN 113234657 A CN113234657 A CN 113234657A
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mercaptoethanol
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CN113234657B (zh
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李向东
周丽君
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China Agricultural University
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Abstract

本发明公开了一种细胞培养用血清替代物，包括如下原料：牛血清白蛋白、胰岛素、转铁蛋白、亚硒酸钠、肝素钠、β‑巯基乙醇、叶酸、乙醇胺、钼酸铵以及微量/少量胎牛血清。本发明的血清替代物，添加到常用的基础培养基里就能使细胞贴壁且增殖，可使成本大幅度降低，降低生产成本。

Description

一种无血清细胞培养基的应用
技术领域
本发明涉及一种培养基用血清替代物，具体涉及一种细胞培养用血清替代物。
背景技术
永生化的细胞系培养经历了几十年的发展，尽管现在大多数细胞系仍然需要用含有血清的培养基培养，但在很多情况下，具有明确组成成分的血清替代物对于后续的科学研究有很多优势：①选择性地促进特殊类型细胞的生长；②调节细胞增殖或是分化的可能性；③降低不同批次的牛血清间的质量差异，从而提高实验的重复性和准确性；④血清替代物较全血清具有明确的成分，避免了组分对实验研究的影响，降低了血清中未知成分的干扰和血源性污染。
但血清替代物的缺点：①不同类型的细胞甚至同一类型的不同细胞(肿瘤细胞)都需要不同的培养基，血清替代物的特异性给实验室保存不同类型的细胞系带来了困难；②血清替代物对试剂、水等的要求会更高。
然而，含有全血清与不含血清都会对细胞的培养产生影响。
发明内容
为了解决现有技术中存在的问题，本发明的目的是提供一种细胞培养用血清替代物，与基础培养基合用可以使细胞贴壁且增殖。
为了实现本发明目的，本发明首先通过以下技术方案实现：
一种细胞培养用血清替代物，包括如下质量浓度的原料：10％-50％牛血清白蛋白、1-10mg/L胰岛素、10-100mg/L转铁蛋白、5-10μg/mL亚硒酸钠、10-200ng/mL肝素钠、20-100μM β-巯基乙醇、1％-10％叶酸、0-100mM乙醇胺、0-100mg/L钼酸铵、0-1％胎牛血清。
一种细胞培养用血清替代物，包括如下质量浓度的原料：10％-50％牛血清白蛋白、1-8mg/mL胰岛素、10-70mg/L转铁蛋白、5-8ug/mL亚硒酸钠、10-100ng/mL肝素钠、20-80μMβ-巯基乙醇、1％-8％叶酸、0-50mM乙醇胺、0-100mg/L钼酸铵、0％-0.5％胎牛血清。
一种细胞培养用血清替代物，包括如下质量浓度的原料：10％-25％牛血清白蛋白、1-6mg/L胰岛素、10-50mg/L转铁蛋白、5-8ug/mL亚硒酸钠、50-100ng/mL肝素钠、20-60μMβ-巯基乙醇、1％-5％叶酸、0-50mM乙醇胺、100mg/L钼酸铵、0％-0.5％胎牛血清。
本发明的有益效果在于：
本发明一种血清替代物的应用，与基础培养基合用就能使细胞贴壁和增殖，完全代替全血清的使用；可使成本大幅度降低，降低生产成本。
附图说明
此处所说明的附图用来提供对本发明实施例的进一步理解，构成本申请的一部分，并不构成对本发明实施例的限定。在附图中：
图1为实施例1中实验组与对照组的培养基培养hela、hepG2、MDA-MB-231、293T细胞形态图；
图2为实施例4中实验组的培养基培养hela、hepG2、MDA-MB-231、293T细胞形态图；
图3为分别采用添加本发明的培养基、添加10％血清的培养基以及基础培养基培养hela、hepG2、MDA-MB-231、293T细胞的细胞增殖能力的结果。
具体实施方式
以下实施例用于说明本发明，但不用来限制本发明的范围。
实施例1
一种细胞培养用血清替代物A，包括如下质量浓度的原料：10％-25％牛血清白蛋白、1-6mg/L胰岛素、10-50mg/L转铁蛋白、5-8ug/mL亚硒酸钠、50-100ng/mL肝素钠、20-60μMβ-巯基乙醇、1％-5％叶酸、0-50mM乙醇胺、100mg/L钼酸铵、1％胎牛血清。
实施例2
一种细胞培养用血清替代物B，包括如下质量浓度的原料：10％-25％牛血清白蛋白、1-6mg/L胰岛素、10-50mg/L转铁蛋白、5-8ug/mL亚硒酸钠、50-100ng/mL肝素钠、20-60μMβ-巯基乙醇、1％-5％叶酸、0-50mM乙醇胺、100mg/L钼酸铵、0.5％胎牛血清。
实施例3
实验组：采用DMEM-highglucose基础培养基，添加本发明实施例1的细胞培养用血清替代物。
对照组1：采用DMEM-highglucose基础培养基，添加10％胎牛血清。
对照组2：DMEM-highglucose基础培养基。
用上述实验组与对照组的培养基培养hela、hepG2、MDA-MB-231、293T细胞(图1)。
实施例4
采用DMEM-highglucose基础培养基，添加本发明实施2的细胞培养用血清替代物。
用上述培养基培养hela、hepG2、MDA-MB-231、293T细胞(图2)，以及增殖倍数结果(图3)。
如图3所示，细胞在实验组和实施例4培养基中的增殖倍数高于对照组2且与对照组1相当，提高细胞培养的质量。
虽然，上文中已经用一般性说明及具体实施方案对本发明作了详尽的描述，但在本发明基础上，可以对之作一些修改或改进，这对本领域技术人员而言是显而易见的。因此，在不偏离本发明精神的基础上所做的这些修改或改进，均属于本发明要求保护的范围。

Claims (3)

1.一种细胞培养用血清替代物，其特征在于，包括如下质量浓度的原料：10％-50％牛血清白蛋白、1-10mg/L胰岛素、10-100mg/L转铁蛋白、5-10μg/mL亚硒酸钠、10-200ng/mL肝素钠、20-100μMβ-巯基乙醇、1％-10％叶酸、0-100mM乙醇胺、0-100mg/L钼酸铵、0-1％胎牛血清。
2.根据权利要求1所述的一种细胞培养用血清替代物，其特征在于，包括如下质量浓度的原料：10％-50％牛血清白蛋白、1-8mg/mL胰岛素、10-70mg/L转铁蛋白、5-8ug/mL亚硒酸钠、10-100ng/mL肝素钠、20-80μMβ-巯基乙醇、1％-8％叶酸、0-50mM乙醇胺、0-100mg/L钼酸铵、0％-0.5％胎牛血清。
3.根据权利要求1所述的一种细胞培养用血清替代物，其特征在于，包括如下质量浓度的原料：10％-25％牛血清白蛋白、1-6mg/L胰岛素、10-50mg/L转铁蛋白、5-8ug/mL亚硒酸钠、50-100ng/mL肝素钠、20-60μMβ-巯基乙醇、1％-5％叶酸、0-50mM乙醇胺、100mg/L钼酸铵、0％-0.5％胎牛血清。
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Cited By (2)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN114214287A (zh) * 2021-12-28 2022-03-22 广州泛恩生物科技有限公司 换液培养基及其应用
CN116574683A (zh) * 2023-07-12 2023-08-11 中国农业大学 一种细胞生长促进剂及其应用

Citations (5)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
JPH0787964A (ja) * 1993-06-03 1995-04-04 K G S:Kk 無血清培養可能な細胞とワクチンウイルスの製造法
CN101984051A (zh) * 2010-11-19 2011-03-09 中山大学 适合肿瘤干细胞富集与培养的无血清细胞培养液
CN103060264A (zh) * 2012-12-20 2013-04-24 上海市第十人民医院 一种干细胞培养基及其应用和干细胞培养方法
US20140178987A1 (en) * 2012-12-20 2014-06-26 Shanghai Tenth People's Hospital Stem cell culture medium and its applications as well as a stem cell culture method
CN105524882A (zh) * 2016-01-18 2016-04-27 华东理工大学 用于免疫杀伤细胞体外扩增的血清替代物组合

Patent Citations (5)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
JPH0787964A (ja) * 1993-06-03 1995-04-04 K G S:Kk 無血清培養可能な細胞とワクチンウイルスの製造法
CN101984051A (zh) * 2010-11-19 2011-03-09 中山大学 适合肿瘤干细胞富集与培养的无血清细胞培养液
CN103060264A (zh) * 2012-12-20 2013-04-24 上海市第十人民医院 一种干细胞培养基及其应用和干细胞培养方法
US20140178987A1 (en) * 2012-12-20 2014-06-26 Shanghai Tenth People's Hospital Stem cell culture medium and its applications as well as a stem cell culture method
CN105524882A (zh) * 2016-01-18 2016-04-27 华东理工大学 用于免疫杀伤细胞体外扩增的血清替代物组合

Non-Patent Citations (1)

* Cited by examiner, † Cited by third party
Title
张松等: "无血清培养基的研究进展", 《江西畜牧兽医杂志》 *

Cited By (4)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN114214287A (zh) * 2021-12-28 2022-03-22 广州泛恩生物科技有限公司 换液培养基及其应用
CN114214287B (zh) * 2021-12-28 2023-11-03 广州泛恩生物科技有限公司 换液培养基及其应用
CN116574683A (zh) * 2023-07-12 2023-08-11 中国农业大学 一种细胞生长促进剂及其应用
CN116574683B (zh) * 2023-07-12 2023-09-29 中国农业大学 一种细胞生长促进剂及其应用

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